Protoplast preparation and induced screening of alkaline pullulanase producing bacterium

The optimal condition for preparating of the protoplast and its regenerating were studied. Under 0.5 mg/mL of lysozyme,the preparation ratio reach to 93.8% when cells were lysed at 37℃ for 1.5 h.The optimum steadyreagent of protoplast regeneration is mannitol.The regeneration ratio is above 26.4%. Protoplast of Bacillus sp SX12 were induced by ultraviolet ray. One high producing strain (SX12 C67) were obtained from process of screening. The activity of pullulanase in strain SX12 C67 increased from 2.42 μ/mL in strain SX12 to 6.87 uμ/mL, about 1.8 times than strain SX12.